Malaria kills more than one million people each year. Most malaria deaths occur in sub-Saharan Africa and most of the victims are children under five. An effective, affordable vaccine could greatly reduce the morbidity, mortality, and economic burden imposed by malaria. This proposal will investigate one approach to such a vaccine. The immunization approach that we will explore is modeled on existing programs for immunization against human adenoviruses and will employ live recombinant derivatives of adenovirus vaccine strains that have been used safely and effectively for over three decades. These vaccines are administered orally, are protective in one dose, and are relatively inexpensive to produce, making them promising for use in resource-poor locations that lack strong health-care infrastructures. Recombinants will express two pre-erythrocytic Plasmodium falciparum antigens that induce immunity in animals: circumsporozoite protein (CSP) and liver stage antigen 3 (LSA-3). These antigens will be produced by a novel system developed in this laboratory for high-level expression of genes inserted into the adenovirus major late transcriptional unit (MLTU recombinants), or in the form of fusions with major viral capsid proteins designed to display malaria antigenic determinants on the surface of the recombinant particles (capsid display recombinants). We have extensive experience with viable recombinants that abundantly express human and canine papillomavirus L1 genes from the MLTU and have prepared capsid display recombinants that bear antigenic epitopes of the P. falciparum CSP and HPV L2 proteins. The L1 protein produced by MLTU recombinants assembles into VLPs that induce antibodies in mice, and capsid display recombinant particles expressing CSP or HPV L2 induce neutralizing antibody to malaria sporozoites and HPV, respectively. We will prepare recombinants expressing CSP and LSA-3 that are analogous to existing L1 MLTU recombinants, and a range of CSP capsid display recombinants with varying structures. The recombinants will be characterized in vitro for antigen production and growth properties and then evaluated in two animal model systems for immunogenicity and protective efficacy. Capsid display mutants will first be examined in mice for their ability to induce antibody and protect against a sporozoite challenge and optimal constructs will be identified. Promising capsid display recombinants identified in mice and MLTU recombinants selected for desirable properties in tissue culture then will be assessed for the ability to induce humoral and cell-mediated immune responses in a permissive primate model. We will also attempt to develop a primate model in which protection from experimental challenge can be determined, and will measure protective efficacy if that effort is successful. These studies will provide information that can be used to prepare live recombinant vaccines that express malaria antigens for use in human trials.